LIGHT-DRIVEN ION TRANSPORT IN BACTERIAL RHODOPSINS

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EXCEED THE SPACE PROVIDED. Bacteriorhodopsin is the simplest of ion pumps, and the structural prototype of the ubiquitous heptahelical membrane proteins, in the last few years we determined the x-ray diffraction structures of seven trapped photointermediates of this light-driven proton transport system. The structures of the remaining two states are available indirectly from non-illuminated mutants. The conformational transformations of the retinal and opsin suggest an atomic level mechanism for this pump, but the mechanism is incomplete without the structures of all nine intermediates. The states with less direct information are those where large-scale motions of the transmembrane helices mediate the entry and exit of buried water molecules that affect the pKs of proton donors and acceptors and form proton conducting chains. These states cannot be trapped in 3-dimensional crystals. We will focus on testing and authenticating the structures of these crucial states of the transport cycle by site-directed spin-labeling. A large number of strategically located cystine residues will reveal how well the mutant structures correspond to the N, N' and O intermediates, and if different, begin to define the strucures of these states. At the same time, crystallographic studies with trapped states will continue to improve the structures already available, and to use visible and infrared spectroscopy to explore still outstanding questions concerning proton transfers in the extracellular region of the protein. The outcome of this work is expected to be the first full description of the coupling of local changes at the active site to global changes of the protein in any transporter or receptor. PERFORMANCE SITE ========================================Section End===========================================